The present invention relates to a cholesterol sensor that facilitates rapid and simplified quantitation of cholesterol and cholesterol ester concentrations contained in a sample with accuracy, and a method for producing the same.
Proposed conventional system for simplified quantitation of specific components contained in a sample without requiring dilution or agitation of sample solution includes a biosensor disclosed in Japanese Laid-Open Patent Publication Hei 2-062952.
The biosensor is produced by the steps of forming an electrode system comprising a measuring electrode, a counter electrode and a reference electrode by screen printing or the like on an electrically insulating base plate, and disposing thereon an enzyme reaction layer containing a hydrophilic polymer, an oxidoreductase, and an electron mediator. The enzyme reaction layer may further contain a buffer, if necessary.
Upon dropping aliquots of a sample solution containing a substrate over the enzyme reaction layer of such biosensor, the enzyme reaction layer dissolves, causing reaction between the enzyme and the substrate, and reduction of the electrode mediator will result. Upon completion of enzyme reaction, the reduced electron mediator is electrochemically oxidized. The concentration of the substrate in the sample solution can be determined by measuring the oxidation current value at that time.
Theoretically, this biosensor can be applied in measuring various substances if an appropriate enzyme of which substrate is the target of measurement is selected. For example, the use of cholesterol oxidase as the oxidoreductase facilitates production of a biosensor for measuring serum cholesterol levels.
However, the serum cholesterol level used as a diagnostic index represents a sum of cholesterol and cholesterol ester concentrations. Cholesterol ester per se can not serve as the substrate for causing oxidation reaction by cholesterol oxidase. Therefore, for the measurement of the serum cholesterol level as a diagnostic index, cholesterol ester should be converted to cholesterol by some means. Cholesterol esterase is a known catalyzing enzyme for this conversion.
However, the use of cholesterol esterase as the catalyst for converting cholesterol ester to cholesterol necessitates a problem of very slow reaction.